596
JUNCINS A-F, SIX NEW BRIARANE DITERPENOIDS FROM THE GORGONIAN JUNCEELLA JUNCEA S. ISMCS, S. C A R ~ ~ E Land Y , Y.KASHMAN+ Sackkr Fanrlty of EW Sciencu, School of Cbemistv, TeI Aviv U n i w i t y , Ramut Aviv 69978, TeI A&, Israd A s s m c r . S i x new diterpenoids, juncins A-F 12-7, possessing the known briarane skeleton have been isolated from a gorgonian collected from the Red Sea. The structures and relative stereochemistry of the juncins were determined by extensive 'H and 13C 1D and 2D nmr spectral analysis.
Briarein A 11)isolated from the gorgonian Briarerrmashtinurn was the first reported example of these novel bicarbocyclic diterpenes (1-5). Diterpenes with the same skeleton as briarein A have been subsequently reported from octocorals belonging to the order Pennatulacea and also from Gorgonacea. Most recently a novel briarane diterpenoid was also isolated from a Mediterranean nudibranch and from its octocoral prey (6). Our investigation of the gorgonianJuncdfajunceu Pallas (Gorgonaceae)has resulted in the isolation and identification of six new briarane diterpenes, 2-7.J . junceu was collected from sites in the entrances to the Gulf of Eilat and Gulf of Suez, Red Sea. The CH,Cl,-soluble extract of the freeze-dried horny coral was separated by repeated chromatographies on Si gel and RP- 18 columns; a total of six compounds have been isolated and identified. These compounds, designated juncins A 121,B 131, C 141, D 151, E {6],and F [7), were isolated in yields of 0.00 1 4 . 0 0 8 % (dry wt). The mass spectra of the juncins, together with their 'H- and 13C-nmr spectra (Tables 1 and 2) suggested the briarane skeleton. Characteristic in the mass spectra of 2-7 were the pairs of peaks differing by 2 amu with a relative intensity of 1:3, suggesting one chlorine atom in each compound. Juncin A [2]had the molecular formula C26H33C1010(hreims, see Experimental), m/z 541/543 (3: 1, [MH]+, cims), and showed a uv absorption at A 220 nm (E 9000) and ir absorptions at v 3520, 1780, 1730, and 1720 cm-', typical for an alcohol, a y lactone, and one or more ester functionalities. Comparison of the 'H-nmr spectrum of 2 with that of 1 (7) (Table 1)and the I3C-nmr spectrum with that ofptilosarcone [8](8) (Table 2) suggested for 2 the same substituted ten-membered ring system as in compounds 1 and 8 and in addition an acetate and an exocyclic epoxide. The positions of the latter two functionalities were unequivocally determined by COSY and difference nOe measurements to be at C- 14 and C- 11, respectively. The resonance line assignments of both protons and carbons of the molecule was assisted by an HMQC (9)experiment that established all C-H connectivities and, hence, also all pairs of geminal protons. The COSY experiment established the following correlations: H-2 to H-3, -4, and Me-15; H-3 to H-4; H-4 to H-6, -16a, -16b; H-6 to H-7, -16a, -16b; H-7 to 8-OH; H-9 to H-10; H-10 to H-20a and Me-15; H-12eq to H-l2ax, -13ax, -13eq, -14; H12ax to H-l3ax, -13eq, -20b; H-13eq to H-l3ax, -14; H-13ax to H-14; H-17 to Me18; H-20a to H-20b. These connectivities established the C- 1(including Me- 15) to C8, C-9 to C-14, and C-17 to C-18 segments ofthe molecule, leaving unconnected the C- 1 to C- 14 and C-8 to C-9 and C- 17 bonds and the manner of the closure of the y-lactone. The suggested briarane diterpenoid structure was unequivocally established by difference nOe measurements summarized in Figure 1. The latter experiment, together with the measured coupling constants (Table 1). also determined the relative
Isaacs et al. : Briarane Diterpenoids
May-Jun 1990)
m 3 e
8 N 0'
n
e
C
E
a
. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
597
598
"C nmr of Juncins A (21, C (47, D (51, and E (61 in CDCI,.'
TABLE 2.
Compound
Carbon
. . . . . . . c-8 . c-9 . c-10 . c-1 c-2 c-3 c-4 c-5 C-6 c-7
c-11 . 6-12 . C-13 . C-14 . C-15 . C-16 . C-17 . c-18 . C-19 . c-20 . OAc's
Wol. 53, No. 3
Journal of Natural Products
. . . . . . . . . .
. . . . . . . . . . .
. . . . . . . . . . .
. .. . .. .
. . . . . . . . .
.
. . .
. . . . . . . . ... . . . . . . . . . . ... . . . . . . .
.
2
4b
47.8s 71.6d 130.5 d 128.2 d 137.0s 62.2d 78.6d 81.3s 70.4d 38.3d 59.3s 25.0td 29.6 td 74.1 d 14.4q 116.9t 47.8d 7.4q 174.9s 50.5 t 21.1 q 21.2q 21.4q
47.6s 70.7 d 130.4d 128.2 d 137.3s 63.8d 78.7 d 80.0s 72.6d' 33.4d 57.9s 72.4 d 29.8 t 72.6d 14.3q 116.8 t 48.4d 8.3q 175.6s 49.7 t 21.lq 21.3q 22.8q
47.7s 70.6d 130.5 d 128.3 d 137.2s 62.2d 78.7 d 80.0s 73.0d' 33.3d 57.8s 72.9d 29.6 t 73.0d' 14.3q 117.0 t 48.0d 8.4q 175.6s 49.5 t 2l.Oq' 21.0q' 21.2q' 21.2q'
169.6s 170.2 se 170.2 se
169.0 s 169.9 s 170.1 s
169.0s 169.7 sr 169.7 sc 170.3 s
6
d
47.1 s 70.8 dd 130.0d 128.5 d 137.2s 64.0d 78.7 d 80.4 s 73.1 dd 32.8d 57.1s 73.5dd 66.8dd 74.7 dd 14.4q 117.lt 48.4d 8.4q 173.1 s 49.5 t 20.5 q 20.7 q 20.9q 21.lq 21.4q 164.3 s 164.4 s 169.5 s 169.7 s 170.3 s
44.72s 76.01 d 131.06d 128.02d 136.56 s 62.91d 72.16d 82.85 s 77.92d 38.86d
5
13.64q 117.79 t 46.28d 7.07q 174.50 s
'Reported as chemical shift, multiplicity. bS13 C alues of the isovalerate: 172.3 s, 43.3 t, 33.4 d, 24.9 q (2C). 'Data from Hendrickson and Cardellina (8). dAssignments are interchangeable. 'Overlapping signals.
stereochemistry of the various c h i d centers, namely, the same 1R*,2S*,6SX,7R*, 8R+,9S*, lose, 1lSx, 12R+,I@*, 17R* configurations as in compounds 1 (1-5) and 8 (8).
The structure for 3, C26H3,CI0,, obtained in minute amounts only, followed directly from the identification of 2. Comparison of the 'H-nmr spectra (Table 1)determined the 11,20-deoxyjuncin A structure for 3. Juncin B is, most likely, the biogenetic precursor of 2. The structure of 3 was confirmed by a COSY experiment which established the following correlations: H-2 to H-3, -4; H-3 to H-4; H-4 to H-6, -16a, -16b;H-6toH-7, -16a;H-9toH-lO;H-l2axtoH-l2eq, -13ax, -13eq, -20b;H-12eq to H-14; H-13ax to H-l3eq, -14; H-l6a to H-16b; H-17 to Me-18; H-20a to H-20b. Juncins C 147,D 151, and E 161all possess the Same ten-membered ring as juncins A 127 and B 131 but differ in the cyclohexane unit and in case of 4 also in one of the four ester groups. Juncin C 147,C31H41C1012(hreims), possesses in addition to three acetates an isovalerate ester, as suggested by the 'H n m r [6 0.99 (d,J=6.2, Me-25), 0.95 (d,
May-Jun 19901
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5 99
0 1 R ~ = A c ,R,=R,=OAc, R4=H 8 R,=C3H,C0, R,, R4=0, R,=H
2 R ~ = R , = R ~ = A cR,=R,=H ,
3 R,=R,=R,=Ac, R4=R,=H, 11,20deoxy 4 R,=Ac, R,, R,, OR4=2 Ac+isovalerate, R,=H 5 R,=R,=R3=Ac, R4=OAc, %=H
6 R,=R,=R3=Ac, R4=R,=OAc 7 3,4 dihydro, R,, R,, R,, OR4=3 Ac+isobutyrate, R,=H
9 R,=R,=H, R,, R,, OR4=2 Ac+isovalerate
FIGURE 1. Difference nOe measurementsof juncin D 157.
J = 6 . 2 , Me-24), 2.10 (m, H-23), 2.06 (m, H-22’), 2.25 (m, H-22)], which was assigned by a COSY experiment (H-22 correlates to H-22’ and H-23 and H-23 to Me-24 and Me-25). The isovalerate was also in full agreement with the [M-C,H,,O,] + (M - 102 amu) fragment observed in the mass spectrum. The location of the fourth ester, at C- 12, was determined by the COSY experiment. Furthermore, a coupling of less than 1 Hz between H- 12 and H-14 due to a W-arrangement determined the 1,3diequatorial configuration for the latter two protons. The latter stereochemistry was confirmed by nOe’s measured between H-14 and Me-15 and between H-12 and H-20a. The position of the isovalerate, however, could not be determined. It has already been shown for the briaran diterpenoids that no nmr differences exist between methines carrying acetate or other esters (lo), thus excluding nmr measurements for the determination of the location of the isovalerate. Mild basic hydrolysis of 4 afforded the 8,9-dihydroxy compound 9. The C-9 position of the newly formed alcohol was established by double irradiations (see Experimental). Under stronger basic conditions or by treatment of 4 with the Z d C u couple ( 11) no selectivity in the hydrolysis of the esters could be observed. The small available amount of juncin C excluded other possible series of reactions. Juncin D {5], C,,H35C10,,, a tetraacetate, possesses almost the same n m r spectrum, apart from the isovalerate, as that of 4 (Tables 1 and 2). A COSY experiment (see Experimental) confirmed the 2,9,12,14-tetraacetate structure for 5. Juncin E 167, C3,H3,C1OI4, mlz 657/659 (3: 1)EMHI+, is apentaacetate (Tables 1
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Wo1. 53, No. 3
and 2). Four of the acetates are the same as in juncin D, and the 6fih was determined to be on C- 13. The equatorial configuration of the latter acetate (J13=, 1% =J12-, 13= 3.3 Hz) was established by nOe enhancements observed on H-3, H- 13, and H- 14while irradiating the Me-15 group. The last compound, juncin F 171, C3,H4,~1OI2,d z 6291631 (3:1), EMHI+, was also isolated in a small amount only. From the H-nmr spectrum (Table 1)it was found that the diene present in compounds 1-6 was replaced in 7 by an exocyclic methylene (C-5=C-16). Furthermore 7 carries three acetates and one isobutyrate 16, 1.16 (d, J = 7 , Me-24), 1.19 (d, J = 7 , Me-23), 2.50(septet, J = 7 , H-22); [M-C4H,02}+ (M+ - 88 mu)]. All four esters were determined to be on the same carbon atoms as in juncins C and D. However, for the same reasons discussed in the case of juncin C, we could not determine the location of the isobutyrate ester. The structure of 7 was confirmed by a COSY experiment showing the following correlations (the numbers with primes point to one out of a pair of methylene protons-the one resonating at higher field): H-2 to H-3', Me-15; H-3 to H-3', -4, -4'; H-3' to H A , -4'; H-4 to H A ' , - 16a, - 16b; H A ' to H- 16b; H-6 to H-7, - 16b; 8-OHto H-9; H-9 to H- 10; H-10 to H20a, Me-15;H-12 toH-13ax, - 1 3 9 , -14;H-13axtoH-l3eq, -14;H-14toMe-l5;H16a to H- 16b; H- 17 to Me- 18; H-20a to H-20b. The suggested stereochemistry of the acetate at C-2 of 7 was based on the vicinal coupling constants (J2,3f = 8.3,J2,3 = 0), which are similar to the ones measured for minabein 3 (J= 7,2) isolated by Ksebati and Schmitz from a Minabae sp. (12). A literature search for similar 11,2O-functionalized compounds disclosed junceellin (13) and junceellin B (14) isolated from the Chinese gorgonian Juncda spamata and praelolide isolated from the Chinese gorgonian Pleraumida praelonga ( 15). The juncins are closely related to the most recently reported junceelloides, isolated from Junceella fragilis ( 16). Worth mentioning are the H-9 to H- 10 coupling constants (Table 1). Cardelha and coworkers (8,lO) already have pointed out the changes of thisJ value with the substituent at C-11. Our findings show that other changes in substitutions of the cyclohexane unit, even if C- 11 remains unchanged, change significantly theJ9, coupling constant, most likely because of changes in the conformation of the ten-membered ring. EXPERIMENTAL Ir spectra were recorded on a Perkin-Elmer Model 177 spectrophotometer. Uv spectra were recorded on a Perkin-Elmer Model 257 spectrophotometer. Low-resolution mass spectra were recorded on a Finnigan402 1 mass spectrometer, source temperature 220-230", pressure of reagent gases for ci spectra: CH4, 0.28 Torr and NH,, 0.15-0.20 Torr; the electron energies in ei mode were 25-35 eV. 'H- and '3C-nmr spectra were recorded on a Bruker AM-360 spectrometer equipped with an Aspect 3000 computer and operating at 360.1 and 90.5 MHz for 'H and I3C, respectively. All chemical shifts are reponed with respect to T M S ( 6 = 0). Hrms were taken on a VG 70 VSEQ instrument. The 2D-nmr experiments were measured on samples in CDCI, at 298°K. The"-" shift correlation experiments were performed with a COSY 45 sequence. The two-dimensional maps were composed of 5 12 X 2K data point spectra. A l-sec recycle delay was allowed between each pulse sequence. Quadrature detection was applied in both dimensions by using the 16-step phase cycling for N-typepeak selection. Data were multiplied with sine bell shaping function, zero-filled to 1K X 2K and then Fourier transformed and symmetrized. For 'H and 13C nmr of the various compounds see Tables 1 and 2.
COLLECTIONAND EXTRACTION.--]. jnncea was collected at depths of 15-25 rn in the entrances to the Gulf of Eilat and the Gulf of Suez, Red Sea. The specimens were frozen immediately after collection. The freeze-dried organism (250 g) was then extracted with CH2C1, to give after evaporation agum (1.1 g). A voucher specimen was kept in our collection in TA-YK 1690. ISOLATION OF THE BMARANE DITIXPENOIDS 2-7.-The crude material was chromatographed on a +-packed column of Merck Kieselgel H . The column was eluted successively with petroleum ether-
May-Jun 19907
Isaacs et ai. : Briarane Diterpenoids
601
EtOAc (8:2 to 3:2) by applying a vacuum to the receiving flask. T h e material was divided into three fractions. Chromatography of the second fraction (100 mg) on an RP-18 column eluted with MeOH-HZO (1:l)gave juncinA(20mg, 0.008%drywt)andjuncinF(3mg,0.001%).Thesondfraction(210rng) was h t chromatographed again on a Kieselgel H column eluted with petroleum ether/EtOAc, and various combined fractions were separately applied to a RP-18 column. Elution of the latter column with H 2 0 with incteasing percentages of MeOH gave juncin C 141 (40 mg, 0.016%), juncin D 151 (15 mg, 0.006%), juncin B 131(3 rng, 0.001%), and juncin E (6)(15 mg, 0.005%). All six compounds were obtained as amorphous powders which could not be crystallized. TICof the new compounds [Si gel eluted with petroleum ether-EtOAc (1: l)] showed for compounds 6 , 5, 4 , 2, 7,and 3 Rfvalues of 0.20, 0.26, 0.36, 0.37, 0.43 and 0.57, respectively. Juncin A @ ] . + I x ] ~ ~ D - ~ ~(c=0.02, " CHCI ; hreirns d z 540.1752 for C26H33C1010(required 3)l. . 540.1762); ir (CHCI,) 3520, 1780, 1730, 1720 crn- ,cims (CH,)dz (rel. int.) [MHl+ 5411543 (22,9), 481/483 (17,4), 421/423 (49,19), 361/363 (54,21), 325 (a), 307 (49), 279 ( 3 4 , 261 (21), 251 (35), 243 (loo), 225 (27). Juncin B [3].+1X]~'D-55~ (e= 0.003, CHCI,); ir (CHCI,) 3540, 1780, 1750, 1730, 1720 cm-'; cims (CH,) m/z (rel. int.) [MH]+ 525/527 (4, l), 491 (18), 465/467 (21,9), 405/407 (100,32), 345/347 (40,14), 329 (26), 309 (33), 281 (19), 205 (16). Juncin C [4].--[~X]'~D-79" (c=0.02, CHCI,); hreirns m/e 640.2304 for C31H41C1012 (required 640.2287); ir(CHC1,) 3520, 1760, 1740, cm-';cims(CH4)m/z(rel. int.)[MH]+ 641/643(11,5), 5811 583(13,8), 521/523(19,7), 461/463(18,6), 443(16), 419(41), 401(31), 383(43), 358(79), 341 (loo), 323 (74), 305 (40), 2 17 (3I), 205 (12), 103 (37). Juncin D [ 5 ] . + ~ ] ~ ~ ~ - 6 8(c=0.007, " CHCI,); ir (CHCI,) 3540, 1770, 17 10 cm-'; cims (CH,) m/z (rel. int.) [MH]+ 599/601 (4,1), 539/541 (3,1), 479/481 (5,2), 419/421 (14,4), 401 (7), 359/361
(23,9), 341 (25), 323 (22). 279 (37), 205 (70), 149 (loo), 99 (22). Junczn E [~].+Ix]~'D -66" (c= 0.02, CHCI,); ir (CHCI,) 3540, 1780, 1750, 1740, 1720 cm-'; cims (CH,) m/z (rel. int.) [MH]+ 6571659 (45,16), 5971599 (77,29), 537 (20), 477 (16), 417 (20), 357 (30), 339 (37), 281 (57), 279 (loo), 205 (77), 149 (59). Jmcin F [71.+IX]25D-70 (c=0.004, CHCI,); ir (CHCI,) 3490, 1790, 1740, 1730 cm-'; cirns (CH,) m/z (rel. int.) [MH]+ 629/631 (21,7), 569/571 (9,2), 5411543 (60,20), 481 (27), 421/423 (60,21), 361/363 (100,41), 343 (43), 325 (43), 307 (21), 279 (26), 205 (34), 149 (50).
9-Deucetyljuncin C [8].-Juncin C (2 mg) dissolved in a solution of 0.1% K2C0, in MeOH (1 ml) was stirred at room temperature for 20 h. The reaction mixture was neutralized with HOAc-MeOH (1: IO). CHCI, (5 rnl) and H,O (5 ml) were added, the two phases were separated, and the organic layer was dried over MgS0,. The residue after evaporation of the solvents afforded 9deacetyl juncin C: C,,H,,CIOll; m/z [MH]+ 599/601 (22,7); 'H nrnr (360) MHt, (CDCI,) 6.26 ( I H , d , j = 10.3, H-2), 5.98 ( l H , d, J = l l . 9 , H - 4 ) , 5 . 7 6 ( 1 H , d d , J = 1 0 . 3 , 11.9,H-3), 5.62(lH,m,H-16),5.51(lH,s,H-16'),5.45 ( l H , t, H-14), 5.10(1H, d,J=1.7,H-6), 5.07(1H, t , J = 3 , H-12),4.84(1H,d,J= 1.7, H-7),4.00 ( l H , s, 9-OH), 3.64 ( l H , m, H-lo), 3.40 ( l H , rn, H-9), 2.90(1H, q , J = 7 , H-17), 2.79(1H, d, ]=3.5, H-2Oa),2.70(1H,~,8-OH),2.41(1H,d,~~3.5,H-20b),2.16(3H,~,OA~),2.00 OAc), 1.42(3H,d,J=7,Me-18), 1.09(3H, d,]=6.4, Me-24), 1 . 0 1 ( 3 H , d , j = 6 . 4 , Me-25). ACKNOWLEDGMENTS
We express our appreciation to Harbor Branch Oceanographic Institution for financial support, Dr. Y. Benayahu for the collection of the gorgonian, Dr. M.Cojocaru for the mass spectra, and Dr. P. Bell for the hrms measurements. LITERATURE CITED 1. J.E. Burks,D.VanderHelm,C.Y. Chang,andL.S.Ciereszko,ActaCrysrallogr.,B33,704(1977). 2. B.N. Ravi, J.F. Marwood, and R.J. Wells, Ausr. J . Chem., 33, 2307 (1980). 3. S.H. Grode, T.R. James, and J.H. Cardellina 11, Tetrahedron Lett., 24, 691 (1983). 4. J.H. Cardellina 11, T.R. James Jr., M.M.H. Chen, and J. Clardy,]. Org. Chem., 4 9 , 3398 (1984). 5 . J.H. Cardellina 11, Pure Appl. Cbem., 98,365 (1986). 6. A. Guerrieno, M. D'Arnbrosio, and F. Pietra, Helv. Chim. Acta.. 70, 984 (1987). 7. S.J. Wratten, W . Fenical, D.J. Faulkner, and J.C. Wekell, Tetrahedron Lett., 18, 1559 (1977). 8. R.L. Hendrickson and J.H. Cardellina 11, Tetrahedron, 23, 6565 (1986). 9. M.F. Summers, L.G. Marzilli, and A. Bax,J. Am. Chem. Sor., 108,4285 (1986). 10. S.H. Grode, T.R. James Jr., J.H. Cardellina 11, and K.D. Onan,J. Org. Chem., 48,5202 (1983). 11. A. Groweiss, S.A. Look, and W . Fenical, J. Org. Cbem., 48,5203 (1983).
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12. M.B.Ksebati and F.J. Schmitz, Bull. Sor. Chim. BJg., 95,835 (1986). 13. J. Yau, J. Qian, H. Fan, K. Shin, S. Huang, Y.Lin, and K. Long, Zbongsban D a w ?heban Ziran Kcweban, 1,83 (1984);C h n . A b . , 101,21149IC (1984). u Ziran Kexneban, 2, 15 (1987);C h . 14. K. Long, Y. Lin, and W.Huang, Zbongshan Daxw W A h . , 107,215049m (1987). 15. Y.Luo, K.Long, and 2.Fang, Zbongshan Daxw Xnebau Ziran Kexucban, 1,83(1983);C h n . Abstr., 99,50572~(1983). 16. J. Shin, M.Park,and W. Fenical, Tetrakdmn, 45, 1633 (1989). Receid 14 August 1989
